The broad long-term objective of this research is to elucidate the molecular mechanisms underlying sensitivity to the Drosophila male-specific volatile pheromone, 11-cis vaccenyl acetate (cVA). This work is significant because understanding pheromone signal transduction in Drosophila will translate into new methods to control reproduction in insects that transmit human diseases and destroy crops. In the last funding cycle, we made significant progress toward elucidating the mechanisms underlying cVA perception. We discovered a unique signaling paradigm in which activation of pheromone-sensitive neurons is mediated by conformational activation of the extracellular odorant binding protein, LUSH. We have identified several of the proteins that are components of the neuronal receptor complex triggered by activated LUSH, including Or67d, a member of the Drosophila odorant receptor family, and SNMP, a Drosophila homolog of CD36, a membrane protein implicated in atherosclerosis and dyslipidemia in humans. We also found four new gene products essential for normal pheromone detection in a forward genetic screen. In the next funding cycle, we are poised to make rapid progress by exploring how these factors mediate cVA sensitivity. We propose a multidisciplinary approach that includes genetics, electrophysiology, biochemistry, cell and structural biology. The first aim is to further define what activated LUSH interacts with on the T1 dendrites in the pheromone detection cascade. We will also initiate experiments to understand how a single amino-acid mutation in Or67d eliminates signaling. The second aim is to establish if the conformational activation mechanism we have uncovered for LUSH is conserved for other OBPs. The third aim is to define the roles of four new cVA sensitivity factors, odd, vans, vainsB and vainsE identified in the forward genetic screen. Aim 4 is to identify the components required for cVA signal detection mapping to the second chromosome, bringing us closer to our overall goal of identifying the complete set of gene products important for cVA sensitivity. The completion of the studies outlined here will advance our understanding of volatile pheromone signaling in insects, will identify new targets to manipulate insect behavior, and will guide future studies on pheromone signaling in more complex animal model systems. PUBLIC HEALTH RELEVANCE: Insects carry human diseases that kill millions of people every year; malaria alone accounts for over one million deaths annually. Insects also impact agriculture, destroying billions of dollars worth of crops annually. Many insect behaviors, including mating, are triggered by perception of pheromones. Understanding how pheromones are detected at the molecular level will provide new targets to manipulate pheromone communication, with the long-term goal of blocking mating signals and other pheromone-guided behaviors in pathogenic insects. We have identified several genes essential for volatile pheromone detection in Drosophila and will elucidate the roles of these factors in pheromone detection.